Received: 8 March 2021 Revised: 7 April 2021 Accepted: 1 May 2021

DOI: 10.1002/ca.23754

ORIGINAL COMMUNICATION

Clinical anatomy considerations on the muscular and vascular

components of the midface by ultrasonographic imaging

Alonso Hormazabal-Peralta1 | Kang-woo Lee1 | Hyung-Jin Lee1 | You-Jin Choi1 |

Kyung-Seok Hu1 | Hee-Jin Kim1,2

1
Division in Anatomy and Developmental
Biology, Department of Oral Biology, Human Abstract
Identification Research Institute, BK21 PLUS The first signs of face aging appear in the midface, so procedures such as botulinum
Project, Yonsei University College of
Dentistry, Seoul, Republic of Korea toxin and filler injections are performed there. However, no guidelines based on clini-
2
Department of Materials Science & cal anatomy describing the muscular and vascular components in vivo have been
Engineering, College of Engineering, Yonsei
published. The aim of this research was to describe the depths of the midface mus-
University, Seoul, South Korea
cles and the locations of vessels using ultrasonographic (US) imaging. US was applied
Correspondence
at 12 landmarks on the midface in 88 volunteers (49 males and 39 females; 19–
Hee-Jin Kim, Department of Oral Biology,
Yonsei University College of Dentistry, 50-1 36 years) to detect sex differences in the depths of muscles and the locations of the
Yonsei-ro office 601, Seodaemun-gu, Seoul,
vessels. The depths of the orbicularis oculi (OOc), levator labii superioris alaeque nasi
03722, South Korea.
Email: hjk776@yuhs.ac (LLSAN), and zygomaticus minor (Zmi) differed significantly with sex at P7
(p = 0.001) and P8 (p = 0.017), P1 (p = 0.028), and P4 (p = 0.035), respectively. The
Funding information
National Research Foundation of Korea, facial artery, facial vein, angular artery, angular vein, and perforator vessels were
Grant/Award Number: NRF-
found at P9, P2 and P10, P1, P1 and P5, and P8, P11 and P12, respectively. The find-
2017R1A2B4003781
ings indicate that the depths of the OOc, LLSAN, and Zmi muscles differ between
the sexes and that the vessels appear at specific landmarks. This information could
help in developing anatomical guidelines for several procedures.

KEYWORDS
botulinum toxin, facial artery, fillers, levator labii superioris alaeque nasi, midface, orbicularis
oculi, ultrasonography, zygomaticus minor

1 | I N T RO DU CT I O N subsequent sagging of soft tissues. The midface is, therefore, an

The midface refers to the central third of the face and is located
between the glabella and the subnasale point (Farolch-Prats & Nome-
Chamorro, 2019; Milutinovic et al., 2014). It includes mimetic muscles
such as the orbicularis oculi (OOc), levator labii superioris (LLS), leva-
tor labii superioris alaeque nasi (LLSAN), zygomaticus minor (Zmi), and
zygomaticus major (Zmj; Cotofana et al., 2015; Levesque & de la
Torre, 2015). These muscles are involved in various facial expressions
and their repetitive use can lead to the emergence of furrows and
wrinkles in the skin.
A protruding zygomatic bone and an abundance of superficial fat
in the midface contribute to a younger-looking face, whereas aging of
the midface is associated with a decrease in fat volume and the
important anatomical area for facial rejuvenation procedures
(Cotofana et al., 2015; Haffner, 2016). Achieving the best clinical out-
comes without undesirable side effects requires accurate anatomical
knowledge of the three-dimensional (3D) locations and depths of the
various midface muscles and vessels and their surrounding structures
(Spiegel & DeRosa, 2005; Surek et al., 2015).
Ultrasonographic (US) imaging is a versatile method for visualizing
muscular components and obtaining real-time information about inner
structures of the body in a simple, safe, comfortable, and rapid man-
ner within a delimited area (Baloch et al., 2018). It is currently used for
facilitating diagnoses and guiding procedures (Li et al., 2014). In addi-
tion, important vascular components such as the facial artery and vein
or perforator arteries can be recognized using the Doppler mode,

Clinical Anatomy. 2021;1–8. wileyonlinelibrary.com/journal/ca © 2021 American Association of Clinical Anatomists 1

2 HORMAZABAL-PERALTA ET AL.

helping to avoid undesirable outcomes such as bruises or vascular

injection. However, US imaging has not been widely applied to the
facial area owing to the scarcity of US anatomical information about
the midfacial area, despite the availability of long-standing classical
anatomical information.
The present study was performed to establish the depths of the
midface muscle components and the locations of vessels using US
imaging, with the aim of providing critical clinical anatomical guide-
lines for improving several clinical procedures.

2 | METHODS

2.1 | Ultrasonography scans

US facial scans of 88 Korean volunteers (49 males and 39 females

aged 19–36 years) were analyzed. Volunteers who had previously
received surgery or esthetic treatment to the face were excluded. The
left side of the face of each volunteer was US-scanned and measure-
ments were taken. All of the US assessments were performed by the
same researcher (K.W.L) at the Yonsei University College of Dentistry
between July 2017 and August 2019.
To locate the components of the midface, four vertical and three
horizontal lines were defined using the inferior orbital rim of the zygo-
matic bone, mid pupillary line, lateral and medial canthi, and nasal
point as references. These lines were used to establish 12 facial land-
marks (P1–P12) that were marked on the faces of the volunteers. The
four vertical lines were defined as passing through the medial canthus
(VL1), pupil (VL2), lateral canthus (VL3), and ateral orbital rim (VL4), F I G U R E 1 Illustration representing the 12 facial landmarks
while the three horizontal lines were defined as passing through the (points) used in this study. VL1: The line passing through the medial
canthus; VL2: The line passing through the pupil; VL3: The line
inferior orbital rim (HL1), midway between HL1 and HL3 (HL2), and
passing through the lateral canthus; VL4: The line passing through the
bilateral to the nose ala (alare point; HL3). The points at which HL1
lateral orbital rim; HL1: The line passing through the inferior orbital
intersected VL1, VL2, VL3, and VL4 were labeled P1, P2, P3, and P4, rim; HL2: The line midway between HL1 and HL3; HL3: The line
respectively; those at which HL2 interacted with VL1, VL2, VL3, and bilateral to the nose ala (alare point) [Color figure can be viewed at
VL4 were labeled P5, P6, P7, and P8; and those at which HL3 inter- wileyonlinelibrary.com]
acted with VL1, VL2, VL3, and VL4 were labeled P9, P10, P11, and
P12, respectively (Figure 1).
After the facial landmarks were defined, US scanning was per- All of the measurements and the purpose of the study were
formed using a real-time two-dimensional B-mode US device (ECUBE explained to each volunteer, who then signed an informed-consent
15, ALPINION Medical Systems, Seoul, Korea) with a high-frequency form. The study was approved by the Institutional Review Board (IRB)
hockey-stick transducer (8–15 MHz; IO8-17T, ALPINION Medical of Yonsei University (IRB no. 2-2017-0023), and all the examinations
Systems). Each volunteer was placed in a semisupine position and the were performed in accordance with the Declaration of Helsinki.
12 facial landmarks were carefully marked on the face. Before the US
scanning, nontoxic US gel (MeditopSono Jelly, Meditop Corporation,
Seoul, Korea) was applied generously; the transducer was then 2.2 | Statistical analysis
suspended in the gel, taking care to not press the face so that defor-
mity or distortion of the soft tissues due to pressure on the skin was Mean ± SD values were used for descriptive analyses. Data normality
precluded. The gel was reapplied for scanning each landmark. was tested separately for each variable using the Shapiro–Wilk nor-
The left side of the face of each volunteer was US-scanned and mality test. Student's t-test was used to identify significant sex differ-
measurements were obtained. After the images were collected, the ences in the depths of the midface muscles. Height, weight, and BMI
muscular and vascular structures were identified and the depths of (according to World Health Organization standards for the Asia Pacific
the muscles from the skin were measured using an image analysis pro- region; World Health Organization Western Pacific Region, 2000)
gram (ImageJ, National Institutes of Health, Bethesda, Maryland). were compared by the Mann–Whitney test. A nonparametric ANOVA
HORMAZABAL-PERALTA ET AL. 3

(the Kruskal–Wallis test) was used for nutritional status. Data were 3 | RE SU LT S
processed using STATA (version 14.0 for Windows, Lakeway Drive,
College Station, Texas), and p < 0.05 was considered significant. The general characteristics of the sample (N = 88) are presented in
Table 1. Forty-nine males (23 ± 2.69 years old) and thirty-nine
females (25 ± 4.55 years old) participated in the study. The height
TABLE 1 General characteristics of the sample (N = 88)
(p < 0.001), weight (p < 0.001), and BMI (p = 0.003) were signifi-
Male (n = 49) Female (n = 39) cantly greater in males than females. However, according to the
Mean (SD) Mean (SD) p-value standards of the World Health Organization (WHO) for the Asia-
Age 23 (2.69) 25 (4.54) 0.087 Pacific population (World Health Organization Western Pacific

Height 1.74 (0.06) 1.63 (0.05) 0.000 Region, 2000), there was no significant sex-related difference in
nutritional status.
Weight 66.94 (9.14 54.38 (7.51) 0.000
BMI 21.98 (2.30) 20.49 (2.34) 0.003
Nutritional status
3.1 | Basic anatomy at the 12 points
Underweight 2 (4.08%) 6 (12.24%) 0.183
Normal weight 31(63.27%) 26 (53.06%)
At P1 from medial to lateral, the nasalis, LLSAN, and OOc muscles can
Overweight 11 (22.45%) 5 (10.20%) be clearly observed below the subcutaneous tissue layer as hypo-
Obese type 1 5 (10.20%) 2 (4.08%) echoic structures. The angular artery (more medially located) and vein
Note: Mann–Whitney test for comparison of all variables, except run at this point (Figure 2(A)).
nutritional status, where non-parametric ANOVA (Kruskal–Wallis test) At P2 the muscles can be observed beneath an irregular hyper-
was used. Age in years; Height in meters; Weight in kilograms; echoic subcutaneous layer. The OOc can be seen as a thick hypo-
BMI = Weight (kg)/height (m2). Nutritional status is defined according to
echoic line crossing the whole US image, while the LLS is observed as
WHO standards (for Asia Pacific region; World Health Organization
Western Pacific Region, 2000): Underweight below 18.5%, normal weight a shorter hypoechoic line below it (Figure 2(B)). The facial artery can
18.5–22.9%, over-weight 23.0–24.9%, and 25.0–29.9% Obese type 1. be observed running superficially to the LLS.

F I G U R E 2 Ultrasonographic images of the midface. (A) Doppler mode (transverse view, 15 MHz hockey-stick transducer) of P1, which is the
intersection between the infraorbital rim and the medial canthus; (B) Doppler mode (transverse view, 15 MHz hockey-stick transducer) of P2,
which is the intersection between the infraorbital rim and the pupil line; (C) B mode (transverse view, 15 MHz hockey-stick transducer) of P3,
which is the intersection between the infraorbital rim and the lateral canthus. (D) B mode (transverse view, 15 MHz hockey-stick transducer) of
P4, which is the intersection between the infraorbital rim and the lateral orbital rim [Color figure can be viewed at wileyonlinelibrary.com]
4 HORMAZABAL-PERALTA ET AL.

F I G U R E 3 Ultrasonography images of the midface. (A) Doppler mode (transverse view, 15 MHz hockey-stick transducer) of P5, which is the
intersection of the line midway between HL1 and HL3 and the medial canthus; (B) B mode (transverse view, 15 MHz hockey-stick transducer) of
P6, which is the intersection of the line midway between HL1 and HL3 and the pupil line; (C) B mode (transverse view, 15 MHz hockey-stick
transducer) of P7, which is the intersection of the line midway between HL1 and HL3 and line lateral canthus. (D) Doppler mode (transverse view,
15 MHz hockey-stick transducer) of P8, which is the intersection of the line midway between HL1 and HL3 and the lateral orbital rim [Color
figure can be viewed at wileyonlinelibrary.com]

At P3 the OOc is clearly located deeper than the previous land-
marks owing to the thicker subcutaneous tissue and skin (Figure 2(C)).
The thickest subcutaneous tissue on the orbital rim reference line
was observed at P4. Therefore, the OOc was also located deeper. The
origin of the ZMi can be observed deep to the OOc muscle
(Figure 2(D)).
At P5 the joint of the nose and the midface can be observed with
a concave form near the center of the US image. From medial to lat-
eral the nasalis, LLSAN and OOc muscles can be observed. The LLS
can be seen below the superficial OOc. At this point the angular artery
can be seen running superficial to the LLSAN muscle (Figure 3(A)).
At P6 the LLS can be observed just above the maxilla and the
OOc superficial to the LLS (Figure 3(B)).
At P7 the lower part of the OOc can be observed; the LLS and
the bony origin of the ZMi can also be found at this landmark
(Figure 3(C)).
At P8 the OOc and the bony origin of the ZMj can be seen under
a thick layer of fat called the superficial malar fat. Beneath the dermis,
perforator vessels can commonly be identified (Figure 3(D)).
In P9 to P12, it is hard to visualize the muscles owing to the over-
lapping muscles and the large amount of superficial fat tissue. How-
ever, the vascular component is clearly noticeable using the
Doppler mode.
P9 is the alar facial crease region. The facial artery can be
observed through the adipose tissue, which is located above the mus-
cles. The nasalis, LLS, and LLSAN muscles can be seen from medial to
lateral, respectively (Figure 4(A)).
At P10 the LLS and ZMi can be observed below a thick layer of
subcutaneous tissue (superficial malar fat pad), and the facial vein can
be seen running at this point (Figure 4(B)).
At P11, both the ZMi and ZMj muscles can be observed and the
perforator arteries can frequently be seen, superficially located
(Figure 4(C)).
At P12, the ZMj muscle can be seen lying under a thick layer of
superficial malar fat. At this point the transverse facial artery
(or perforator a.) can also be observed superficially, just below the skin
(Figure 4(D)).
Table 2 presents the measured depths of the muscles and com-
parisons between the sexes. The depths of the OOc differed signifi-
cantly at P7 (p = 0.001), 3.5 ± 0.42 mm in males and 3.1 ± 0.47 mm in
females. The depth of the Zmi differed significantly with sex at P4
(p = 0.035), 6.7 ± 0.83 mm in males and 7.3 ± 1.01 mm in females.
Finally, the depth of the LLSAN differed significantly with sex at P1
(p = 0.026), 3.9 ± 0.77 mm in males and 4.4 ± 1.02 mm in females
(Figure 5). No significant sex-related difference was found for the
nasalis, ZMj, or LLS.
HORMAZABAL-PERALTA ET AL. 5

F I G U R E 4 Ultrasonographic images of the midface. (A) Doppler mode (transverse view, 15 MHz hockey-stick transducer) of P9, which is the
intersection of the line bilateral to the nose ala and the medial canthus; (B) Doppler mode (transverse view, 15 MHz hockey-stick transducer) of
P10, which is the intersection of the line bilateral to the nose ala and the pupil line; (C) Doppler mode (transverse view, 15 MHz hockey-stick
transducer) of P11, which is the intersection of the line bilateral to the nose ala and the lateral canthus. (D) Doppler mode (transverse view,
15 MHz hockey-stick transducer) of P12, which is the intersection of the line bilateral to the nose ala and the lateral orbital rim [Color figure can
be viewed at wileyonlinelibrary.com]

4 | DISCUSSION
Several previous studies have investigated the occurrence and distri-
bution of the midface muscles, but few have considered the depths of
the midface muscular components. Therefore, the aim of the current
study was to use US imaging to provide clinical anatomical informa-
tion about the depths of muscular components and the locations of
vascular components in the midface in order to improve several clini-
cal procedures.
We observed that the same muscles could be located using the
same facial landmarks irrespective of sex, but at different depths
(Figure 5).
The Zmi was significantly deeper in females in the present study,
as also reported by Spiegel and deRosa for Caucasian cadaveric speci-
mens (Spiegel & DeRosa, 2005). However, the Asian face differs from
those in other populations owing to its greater amount of malar fat
and its thicker skin and ligamentous components (Ryu et al., 2018;
Shirakabe et al., 2003).
We found that all the muscles tended to be located deeper from
medial to lateral and from upper to lower, which is consistent with the
data reported by Kim et al. (2019), who analyzed Korean and Thai
cadaveric specimens using the Morpheus three-dimensional
(3D) scanning system. Those authors found that both the skin and
superficial fat (subcutaneous tissue) tended to become thicker from
upper to lower and medial to lateral directions in the infraorbital area.
As the muscles lie behind the subcutaneous tissue, their increasing
depths can be related to the progressive thickening of the superficial
fat pads from medial to lateral and upper to lower aspects.
The first signs of facial aging appear in the midface, so many pro-
cedures to combat aging are performed in this area. Since facial reju-
venation procedures are generally performed without direct
visualization, knowledge of the 3D structure of the midface (the loca-
tions and depths of muscles and their relationships with neighboring
structures) is crucial. Most clinicians rely on the palpation of muscles
or observations of the rhytids when they perform procedures such as
botulinum toxin and filler injections; this is inadequate. As seen in the
present study and consistent with previous research (Hur et al., 2018;
Kaplan, 2017), the same muscle can occur at different depths along its
path from origin to insertion, owing for example to anatomical alter-
ations that could have occurred during previous procedures or to sex
differences.
In addition, important vessels such as the facial artery commonly
appear in the midface. Therefore, basing a procedure only on visual
observations and palpation could be dangerous owing to the risk of
6 HORMAZABAL-PERALTA ET AL.

TABLE 2 Comparison of the depths of the muscular components of the midface by sex (N = 88)

All sample (N = 88)

Male (n = 49) Female (n = 39)

P Muscle n Mean (SD) n Mean (SD) p

1 Orbicularis oculi 46 2.6 (0.5) 38 2.4 (0.6) 0.246

Levator labii superioris alaeque nasi 38 3.9 (0.8) 35 4.4 (1.0) 0.028
Nasalis 40 2.6 (0.5) 29 2.6 (0.6) 0.686
2 Orbicularis oculi 49 2.5 (0.5) 39 2.5 (0.6) 0.390
Levator labii superioris 23 4.9 (0.8) 18 5.2 (0.7) 0.218
3 Orbicularis oculi 48 3.1 (0.6) 36 3.0 (0.6) 0.376
4 Orbicularis oculi 49 4.1 (0.7) 39 4.2 (0.6) 0.436
Zygomaticus minor 36 6.7 (0.8) 13 7.3 (1.0) 0.035
5 Orbicularis oculi 48 3.1 (0.4) 39 2.9 (0.5) 0.130
Levator labii superioris alaeque nasi 47 5.0 (0.7) 32 4.9 (0.9) 0.323
Levator labii superioris 36 6.9 (1.0) 26 7.3 (1.0) 0.190
Nasalis 10 2.9 (0.5) 7 3.1 (0.9) 0.688
6 Orbicularis oculi 44 3.3 (0.5) 35 3.2 (0.4) 0.266
Levator labii superioris 39 7.2 (0.9) 27 7.6 (1.0) 0.056
7 Orbicularis oculi 49 3.5 (0.4) 39 3.1(0.5) 0.001
Zygomaticus minor 36 7.3 (0.8) 16 7.2 (0.7) 0.767
8 Orbicularis oculi 31 5.4 (0.9) 14 6.0 (0.6) 0.017
Zygomaticus major 34 8.1 (0.9) 15 8.3 (0.2) 0.682

Note: Student's t-test was used to identify significant sex differences in the depths of the midface muscles; p < 0.05 was considered significant; mean and standard
deviation (SD) in millimeters.
Abbreviation: P, Facial landmark point.

F I G U R E 5 Illustration representing the depth of the midface muscular component by each facial landmark. The right side corresponds to the
male sample (n = 49) and the left side to the female sample (n = 39) [Color figure can be viewed at wileyonlinelibrary.com]
HORMAZABAL-PERALTA ET AL.
targeting the wrong muscle or, even more seriously, intravascular
injection.
We found that the OOc was the most superficial muscle in all
the landmarks where it appeared, ranging from 2.5 to 5.4 mm in
males and from 2.4 to 6.0 in females. Therefore, when the target
muscle is the OOc, a shallow injection is required, especially for pro-
cedures on the landmarks located in the orbital rim reference line,
where the most superficial depths were registered. We observed
that the Zmi lay around 3–4 mm under the OOc, and the ZMj was
about 3 mm deeper than the OOc at all the landmarks where they
were seen together. A deep injection is, therefore, not necessary and
could lead to undesired outcomes. Moreover, a deep injection can
target the Zmi instead of the OOc in the uppermost lateral land-
marks, which could lead to undesired outcomes such as changes
when smiling (Kaplan, 2017).
This study used US to explore the midface muscles and obtain
information in a rapid and noninvasive manner. Previous studies on
different regions of the face, such as the forehead region, demon-
strated the suitability of US for visualizing and investigating facial
muscles, yielding results immediately and thus allowing a procedure to
be monitored and guided while it is being performed (Ahn et al., 2020;
Choi et al., 2019). However, owing to the layered arrangement of the
midface muscles, the thickness of the superficial fat pads, and
the absence of a surrounding fascia, it is difficult to identify the
boundaries among the facial muscles through US images. The clini-
cian's anatomical knowledge and clinical expertise are therefore
required. In addition, the quality of the US images depends on the
device used (including the transducer) and the anatomical knowledge
of the operator. High-frequency hockey-stick US transducers provide
high-resolution images of superficial components, such as the thin
and small midface muscles and superficial soft tissues, and the vascu-
lar component, which was noticed at several landmarks. Hence, these
transducers are becoming ideal options for guiding injections into
small irregular surfaces owing to their smaller footprints (Baloch
et al., 2018).
US is recommended for performing safe and minimally invasive
diagnoses and procedures for esthetic and dermatological purposes
(Iyengar et al., 2018). However, for US examination or US-guided
research, a certain amount of pressure is needed to visualize struc-
tures, which leads to distortion of the soft tissues caused by the direct
contact of the transducer with the skin surface. In the present
research, in order to preclude deformation or distortion of the midface
region's soft tissue characteristics due to pressure from the trans-
ducer, we applied a generous amount of gel, obtaining clear images
with minimal distortion because the transducer did not touch the skin
directly.
Weight gain or higher fat proportions could influence the depths
of the muscles through the expansion of facial fat pads (Wan
et al., 2013). In the present investigation, this aspect was perhaps
reflected in the large standard deviation for each measurement.
Thiemann et al. (2017) reported that in a German population, the soft
fascial tissues increased as BMI increased, mainly in the orbital and
cheek areas. Also, Tower et al. (2020) reported that superficial fat
7
volumes are influenced by sex and BMI, females having greater
amounts of superficial fat than males with similar BMIs and ages.
Most of the individuals included in the present study had a normal
BMI according to WHO standards (World Health Organization West-
ern Pacific Region, 2000), while the muscles were located deeper in
females. However, more specific studies are required to reveal the
real relationships among BMI, fat mass, and fat distribution in the
superficial facial fat pads in Koreans.
The present study was subject to several limitations. First,
data were obtained only from adults living in Seoul, South Korea,
so the results are not necessarily applicable to the entire Korean
population. Second, as the present study was carried out on
healthy young volunteers (mean age 23.8 years), it represents only
part of the normal anatomy of the midface, focused on the
depths of the muscles and locations of the vessels. It would be
interesting to perform similar investigations on other areas and
with other age brackets to describe the changes in anatomy over
the lifespan.
OR CID
Kyung-Seok Hu https://orcid.org/0000-0002-9048-3805
Hee-Jin Kim https://orcid.org/0000-0002-1139-6261
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How to cite this article: Hormazabal-Peralta, A., Lee, K., Lee,
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